1. Field of the Invention
The present invention is directed to DNA molecules responsible for encoding the multi-functional proteins which direct the synthesis of the polyketide platenolide. The present invention also is directed to use of that DNA to produce compounds exhibiting antibiotic activity based on the platenolide structure, including specifically spiramycin and spiramycin analogues and derivatives.
2. Description of Related Art
Spiramycin is a macrolide antibiotic useful in both veterinary and human medicine produced by streptomycetes such as Streptomyces ambofaciens (ATCC 15154). Spiramycin is a 16-membered cyclic lactone, platenolide, with three attached sugar residues. Spiramycin's antibiotic activity is believed to be due to its inhibition of protein synthesis by a mechanism that involves binding of the antibiotic to a ribosome. Spiramycin is structurally similar to another antibiotic, tylosin, and the biosynthetic pathways of both are known to be similar.
The biosynthesis of tylosin has been thoroughly investigated (Baltz et al., Antimicrobial Agents and Chemotherapy, 20(2):214-225(1981); Beckmann et al., Genetics and Molecular Biology of Industrial Microorganisms, (1989):176-186). Polyketides are synthesized via a common mechanistic scheme thought to be related to fatty acid synthesis. The cyclic lactone framework is prepared by a series of condensations involving small carboxylic acid residues. Modifications of the structure, such as ketoreduction, dehydration and enolylreduction, also occur during the processing. The synthesis is driven by a set of large multi-functional polypeptides, referred to as polyketide syntheses.
PCT Publication WO 93/13663 describes the organization of the gene encoding the polyketide synthase of Saccharapolyspora erythraea. The gene is organized in modules, with each module effecting one condensation step. The precise sequence of chain growth and the processing of the growing chain is determined by the genetic information in each module. This PCT application describes an approach for synthesizing novel polyketide structures by manipulating in several ways the DNA governing the biosynthesis of the cyclic lactone framework. In order to adapt this methodology to other polyketides, however, the DNA molecules directing the biosynthetic processing must first be isolated.
The present invention is directed to the DNA sequence for the gene cluster responsible for encoding platenolide synthase, the building machinery of platenolide which is the basic building block of spiramycin. As a result, the present invention provides the information needed to synthesize novel spiramycin-related polyketides based on platenolide, arising from modifications of this DNA sequence designed to change the number and type of carboxylic acids incorporated into the growing polyketide chain and to change the kind of post-condensation processing that is conducted.